1. Field of the Invention
The present invention relates generally to the fields of biochemical reaction and biomedical physics. More specifically, the present invention relates to methods/devices used for enhancing catalytic chemical reaction by adding energy to the reactants therefore to increase the frequency at which reactants reach transition states of reaction.
2. Description of the Related Art
Activation Energy and Reaction Rates: In any given chemical reaction, the equilibrium of the reaction is given by the difference in xcex94G0 for the reaction. The equilibrium concentrations of substrate (A) and product (C) are determined by their difference in free-energy content,
xcex94G0=xe2x88x92RTlnkeq
Wherein R=gas constant, 8.3 JKxe2x88x92molxe2x88x921, T=absolute temperature and Keq=equilibrium constant for the reaction (see FIG. 1).
Heat, when added to a reaction, will alter the free energy content of the reaction and therefore will shift the equilibrium of the reaction. It has also been shown that electromagnetic energy, by translating absorbed energy into translational motion, can enhance chemical reactions. However, there is an activation energy barrier between the substrate and product which is given by xcex94Gxe2x80x2. This xcex94Gxe2x80x2 represents the change in free energy that must be put into the system to reach the transition state (FIG. 1). Similarly, the reaction rate is affected by energy of activation, Ea, that reflects the amount of energy that must be added to a reaction for the reactants to reach the transition state. The Arrhenius rate equation describes this reaction rate and is given by:
k=Aexp(xe2x88x92Ea/RT)
Wherein A=pre-exponential factor and Ea=activation energy. Given then, that ln(k)=ln(A)xe2x88x92Ea/RT, a plot of ln(k) vs 1/T, gives intercept A and slope xe2x88x92Ea/R.
If Ea is high, only a portion of the molecular encounters are energetic enough to result in reaction, but if it is low, a high proportion can react, and the rate coefficient is large. Thus, if the activation energy can be lowered in some way, the reaction proceeds more rapidly.
Anything that stabilizes the transition state relative to reactants will decrease the free energy of activation and therefore increase the reaction rate (FIG. 2). A catalyst lowers the activation energy of the rate determining steps, thereby speeding up the reaction. The role of the catalyst is to permit the formation of a transition state of lower energy (higher stability relative to reactants) than that for non-catalyzed reactions. The catalyst itself does not participate in the reaction stoichiometry (not consumed) and cannot affect the equilibrium position of the reaction. Pauling expressed that stabilization of the transition state of a reaction by an enzyme suggests that the enzyme has a higher affinity for the transition state than it does for the substrate or products. xcex94Gxe2x80x2 therefore, is reduced during catalysis.
Improved PCR: The polymerase chain reaction is a technique used for in vitro and in situ amplification of specific DNA sequences. The process goes in receptive cycles: denaturing, wherein the DNA of interest is denatured for about 4 minutes at 94xc2x0 C.; annealing, wherein the appropriate part of the DNA strands are annealed to the primers (i.e., the antisense DNA fragment of interest) at 50xc2x0 C. for about 2 minutes; and extension, wherein a heat stable enzyme called Taq-DNA-polymerase (Taq) polymerizes the individual DNA bases (deoxyribonucleotides) for 3 minutes, at 72xc2x0 C. Such cycle is repeated for N times (xcx9c20-30 times) with more primers and nucleotides added. For the following cycles, heating parameters may be modified slightly (for example, denaturing for 1 min. at 94xc2x0 C.; annealing for 2 min. at 50xc2x0 C.; and extending for 3.5 min at 72xc2x0 C.). Additionally, the heating parameters for the last cycle are also typically different, for example, denaturing for 1 min. at 94xc2x0 C.; annealing for 2 min. at 50xc2x0 C.; and extending for 10 min at 72xc2x0 C. As a result, the DNA of interest is amplified by 2N.
For the heating protocol example above, the total time for denaturation is 4+(20xc3x971)+1=25 minutes with additional time needed for cooling (if N=20). If it were possible to denature and cool the DNA more efficiently, a significant timesaving would result. Furthermore, the part of the PCR cycle that involves the highest temperatures would be eliminated, and thermal breakdown of the reaction buffer materials would be reduced. Further, increasing the reaction rate of the annealing and extension phases of PCR would also add a timesaving.
Biochemical Reaction Catalysis: In order to undergo a chemical reaction, a reactant or reactants must first gain energy (activation energy) to form an activated complex before they can proceed forward to a state (products) of different energy or enthalpy. Generally, enzymes are used to help the reaction take place (i.e. catalyze the reaction). Heat can also be used to speed up the reaction rate. Additionally, vibrational excitation can be used to promote endoergic (energy consuming) reactions [1].
The prior art is deficient in the lack of effective means of enhancing reaction catalysis by adding energy to the reactants. The present invention fulfills this long-standing need and desire in the art.
The present invention provides methods of enhancing reaction catalysis by adding energy to reactants. Specifically, the present invention provides a method of using electromagnetic energy or propagating pressure waves to enhance reaction, wherein vibrations, rotations, and/or particular configurations or orientations of the reactants are induced. When done properly, these methods do not cause irreversible damage to the reactants and are especially suitable for use in biochemical reactions both in vitro and in vivo.
In one embodiment of the present invention, there is provided a method of accelerating a chemical reaction by applying electromagnetic or mechanical energy to the reaction mixture. Preferably, the electromagnetic energy is generated by a source which provides radiant energy with wavelength from about 200 nm to about 20,000 nm. Representative examples of electromagnetic energy include radiofrequency wave and microwave, and representative example of mechanical energy is a pressure wave. Still preferably, the chemical reaction is a catalyzed reaction, it can be either a liquid-phase reaction or a solid-phase reaction.
In another embodiment of the present invention, there is provided a device for accelerating a chemical reaction, comprising a reaction chamber; and a means for applying energy to the reaction chamber.
In still another embodiment of the present invention, there is provided a method of enhancing a polymerase chain reaction by applying energy to the reaction. Such method can also be used for enhancing an enzyme linked immunoassay reaction.
In yet another embodiment of the present invention, there is provided a device for enhancing a polymerase chain reaction, comprising a reaction chamber; and a means for applying energy to the reaction chamber. Such device can also be used for enhancing an enzyme linked immunoassay reaction.
In still yet another embodiment of the present invention, there is provided a method of increasing the rate at which a group of molecules reaches a different molecular configuration from initial configuration, comprising the step of applying energy to the molecules. Preferably, the energy is electromagnetic energy or mechanical energy. Representative examples of electromagnetic energy include radiofrequency wave and microwave, and representative example of mechanical energy is a pressure wave. Still preferably, the different molecular configuration is a transition state in a chemical reaction, preferably, a catalyzed chemical reaction.
In yet another embodiment of the present invention, there is provided a device for increasing the rate at which a group of molecules reaches a different molecular configuration from initial configuration, comprising a chamber for holding the molecules; and a means for applying energy to the chamber.
Other and further aspects, features, and advantages of the present invention will be apparent from the following description of the presently preferred embodiments of the invention given for the purpose of disclosure.